Acta Biochimica et Biophysica 13. (1978)

1978 / 4. szám - Alkonyi, I.-Gyócsi, L.-Sümegi, B.: Demonstration of a Lag Period in the Time-Course of the Reaction Catalyzed by Pyruvate Dehydrogenase Complex

Acta Biochim. et Biophys. Acad. Sei. Hung. Vol. 18 (4), pp. 253— 258 (1978) Demonstration of a Lag Period in the Time-Course of the Reaction Catalyzed by Pyruvate Dehydrogenase Complex I. Alkonyi, L. Gyócsi, В. Sümegi Institute of Biochemistry, University Medical School, Pécs, Hungary (Received April 13, 1978) At low thiamine pyrophosphate concentrations the time-course of the reaction catalyzed by mammalian pyruvate dehydrogenase complex shows a lag period of some minutes when the reaction is started by either enzyme,'pyruvate or thiamine pyrophos­phate. However, started by Co ASH or NAD+, the lag period disappears. An increase in enzyme concentration to 25 mU/ml causes a concomitant short­ening of the duration of the lag period (r), while above this value r is independent of the enzyme concentration. An increase in thiamine pyrophosphate concentration decreases the value of r and the lag period vanishes at infinite thiamine pyrophosphate concentration. It is suggested that both isomerization* and aggregation-dissociation reactions may play an important role in the development of the lag period of pyruvate dehydro­genase complex. Introduction Of the three components of mammalian pyruvate dehydrogenase complex (PDC) the pyruvate dehydrogenase component (Ej) seems to have an outstanding role in the regulation of the enzyme activity. The site of regulation by the phos­phorylation-dephosphorylation cycle is located in the Ex component (Reed et al., 1970, 1972; Hucho, 1975). Pyruvate dehydrogenase exists as a tetramer, <x2ß2, which can bind two molecules of thiamine pyrophosphate (TPP). The available values of Km for TPP are inconsistent (Cooper et al., 1974; Walsh et al., 1976). Investigating the enzyme reaction at low TPP concentrations we have ob­served a transient phase (lag period) lasting for minutes. This phenomenon has not been described, probably because until recently methodological difficulties con­cerning the removal of TPP from the enzyme complex have not been overcome. Transient phases lasting for as long as minutes have been also described (Frieden, 1970; Kurganov et ah, 1976). A more detailed study of this phenomenon shows this transient phase to depend on the manner of the initiation of the reaction. It disappears when the enzyme complex is preincubated in the presence of both pyruvate and TPP. * Also referred to as “conformational transition” and “conformational change” (Kurganov et al., 1976). Acta Biochimica et Biophysica Academiae Scientiarum Hungaricae 13, 1978

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